Hypoxia is a well-characterized component of the solid tumor microenvironment that promotes resistance to radiation therapy and chemotherapy and is associated with a poorer overall prognosis. The hypoxic tumor cell elicits both HIF-dependent and HIF-independent mechanisms to adapt to and overcome the stress of low oxygen in tumors. Through experiments performed under the specific aims of the previously funded application and preliminary results in this proposal, we have shown that hypoxia/anoxia rapidly activates a translational control program mediated by activation of the endoplasmic reticulum (ER) kinase PERK and phosphorylation of the translation initiation factor elF2?. This pathway downregulates global protein synthesis but at the same time upregulates the expression of select stress-response proteins. These results together with those from other groups established that the Unfolded Protein Response, a cellular adaptation mechanism to ER stress, is activated by hypoxia and is required for hypoxia tolerance and tumor growth. In this application we propose to expand these findings to identify the signal for UPR activation by hypoxia, identify the role(s) of downstream effectors of PERK and elF2? phosphorylation, characterize pro- and anti- apoptotic pathways activated by ER stress and test whether the reliance of hypoxic tumor cells on UPR activation for survival can be therapeutically exploited. In Specific Aim 1 we will test whether lack of the downstream PERK effector ATF4 expression inhibits, and overexpression of ATF4 promotes, tumor growth and we will identify and characterize transcriptional targets of hypoxia-activated ATF4. Under Specific Aim 2 we will attempt to delineate the mechanism responsible for PERK and UPR activation under hypoxia by analyzing the status of-SH groups on the ER-resident folding enzymes Protein Disulfide Isomerase, and Erolp oxidase. In Specific Aim 3 we will investigate the role(s) of ER-targeted bcl-2 and ER-localized bax/bak in the induction of ER-dependent apoptosis by hypoxia/anoxia in UPR-proficient and -deficient cells. Under Specific Aim 4 we will test whether hypoxic tumor cells, which are resistant to genotoxic chemotherapeutic agents, are acutely sensitive to pharmacological ER stressors and whether the combination of ER stressors with ionizing radiation or chemotherapeutic agents results in better inhibition of tumor growth in animal tumor models. Solid tumor metastasis and resistance to most established chemotherapy regimens are key contributors to tumor morbidity and thus constitute a significant public health problem. Successful completion of the proposed studies will uncover a novel pathway that contributes to tumorigenesis and resistance to therapy and can potentially offer new targets and approaches to selectively attack these resistant cells and thus improve therapeutic outcome.